The goal of the proposed research is to produce human monoclonal antibodies with specificity for influenza virus antigens and to define their immunologic and biologic properties. Peripheral blood B lymphocytes from persons with recent exposure to influenza antigens will be infected with Epstein-Barr virus to produce continuous lymphoblastoid cell lines. Tissue culture fluids will be screened for influenza antibody by an ELISA technique that utilizes purified influenza virus and enzyme conjugated antihuman IgG antibodies. Secreting cell lines will be grown in cloning conditions and the antibody will be harvested at all stages of growth. Antibody specificity will be tested by ELISA, hemagglutination-inhibition and microneutralization tests using strains representative of major variants within a subtype. They will also be tested for their ability to inhibit binding of representative mouse monoclonal antibodies to each of the four proposed antigenic sites on the hemagglutinin molecule. Comparisons with mouse antibodies will determine if both species identify similar antigenic structures on the viral hemagglutinin. Such information is essential for interpreting the relevance of studies of antigenic variation currently performed with the mouse reagents. This antigenic variation, or "drift," of influenza virus is an important factor in the yearly outbreaks of human disease. The relative avidity/affinity of antihemagglutinin human monoclonal antibodies will be determined by competition radioimmunoprecipitation assays and, in combination with the antibody concentration, and IgG subclass, will be used to compare the efficiencies with which the antibodies inhibit viral hemagglutination and neutralize virus infectivity. This is important for understanding the biologic tests used in assessments of immunity to influenza and may provide information on vaccine formulation so that optimal protective responses will be induced. Possible antigenic differences between egg-grown and tissue culture-grown virus will be sought with human monoclonal antibodies. In the future human monoclonal antibodies may serve as laboratory reagents for analysis of influenza strains or standardization of serological tests and eventually they may be candidates for human therapeutic use.